At present, “abnormal structural proteins” have drawn attention as common mechanisms of developing a plurality of neurodegenerative diseases that develop with aging, such as Alzheimer's disease, Parkinson's disease, Huntington's chorea, and prion disease, and molecular nature of such proteins has been studied. Deposition of the following two types of fibrillar aggregates in the brain has been reported as the pathological feature of Alzheimer's disease: senile plaque primarily composed of amyloid β-proteins (Aβ) (see Selkoe, D. J., Annu Rev. Neurosci., 12, 463-490, (1989); and Glenner, G. G. and Wong, C. W, Biochem. Biophys. Res. Commun., 120 (3), 885-890, (1984)); and neurofibrillary tangles (paired helical filament (PHF)) primarily composed of phosphorylated tau-proteins (see Ihara, Y. et al., J. Biochem., 99, 1807-1810, (1986); and Grundke-Iqbal, I. et al., Proc. Natl. Acad. Sci. U.S.A., 83, 4913-4917, (1986)). As to Alzheimer's disease, which has been considered being caused by a plurality of various pathogeneses, it is now considered through recent studies that the aggregation of amyloid β-proteins should be a common pathway for the development of all such pathogeneses. Amyloid β-protein is a peptide that is cleaved as a molecular species consisting of 40 (Aβ1-40) or 42 (Aβ1-42) residues from its precursor substance (i.e., amyloid precursor proteins (APP)), and the processes of generation and decomposition of amyloid β-proteins as monomers advance homeostatically even in normal humans. In Alzheimer's disease, however, amyloid β-proteins aggregate, and excessive deposition of amyloid β-proteins is observed in the end. This is considered to result from dysregulation during cleavage or decomposition. In the present specification, the former proteins (Aβ1-40) are referred to as “amyloid β40”, “amyloid β40 monomers”, or “monomeric amyloid β40-proteins” in some cases, and the latter proteins (Aβ1-42) are referred to as “amyloid β42”, “amyloid β42 monomers”, or “monomeric amyloid β42-proteins” in some cases. The amyloid β-proteins are cleaved as a molecular species consisting of 43 (Aβ1-43) residues, though in minute quantities, and such proteins may be referred to as “amyloid β43”, “amyloid β43 monomers”, or “monomeric amyloid β43-proteins” in some cases.
The amyloid β-proteins having aggregated act on neurons as neurotoxins and cause synaptic degeneration and subsequent neuronal cell death. This mechanism is considered to cause neuronal loss, which may cause progressive cognitive disorder of Alzheimer's disease. Also, it has been reported that amyloid β-proteins do not exhibit neuronal cell death activity when they were released extracellularly as water-soluble monomeric peptides (hereinafter in the present specification the term “neuronal cell death activity” may be referred to as “toxicity”) and that amyloid β-proteins self-assemble and form amyloid β fibrils, upon which they acquire toxicity (see Lorenzo, A. and Yankner, B. A., Proc. Natl. Acad. Sci. U.S.A., 91, 12243-12247, (1994)). Since it is known that cultured neurons are led to death when a solution containing toxic amyloid β-proteins that contains amyloid β fibrils is added at a high concentration thereto, the amyloid β fibrils have been considered to be the entity to induce neuronal cell death in Alzheimer's disease.
Thus, an experimental system that adds toxic amyloid β-proteins containing amyloid fibrils to neuronal cells and the like so as to induce death of these cells has been considered to reflect the neuronal cell death in Alzheimer's disease and has often been employed in screening inhibitors of neuronal cell death or the like. In recent years, however, the following facts have been reported, which would suggest that the toxic entity of the amyloid β-proteins is not the amyloid β fibrils. That is, (1) the concentration of amyloid β fibrils in a toxic amyloid β-protein-containing solution necessary for inducing neuronal cell death is several tens of μM (see Yankner, B. A., et al., Science, 250, 279-282, (1990)), which is 1,000 times or greater than the concentration of amyloid β-proteins in the brains of patients with Alzheimer's disease; (2) the amount of amyloid β fibrils deposited in the brains of patients with Alzheimer's disease is not always correlated with the impairment of higher-order functions, such as memory or cognitive function, and no clinical symptom may be developed in some cases even though a large amount of amyloid β fibrils are deposited; (3) the site of amyloid β deposition is not always consistent with the site of neuronal drop out in the brain; (4) abnormality is observed in learned behavior before or without the deposition of amyloid β fibrils in the brains of APP-overexpressing mice; and (5) increase in the water-soluble amyloid β-protein content in the brains of patients of Alzheimer's disease occurs 10 or more years ahead of the deposition of water-insoluble fibrils.
The present inventors had proposed a solution containing highly toxic self-assembling amyloid β-proteins that would induce neuronal cell death at a concentration equivalent to that of the amyloid β-proteins that have self-assembled and that exist in the bodies of patients of Alzheimer's disease or other diseases, and had proposed a method for producing such a solution (JP 2001-247600 A). The present inventors had also discovered a method for isolating a neurotoxic entity contained in the aforementioned solution containing self-assembling amyloid β-proteins, and analyzed the same. As a result, such neurotoxins were found to be self-assembling amyloid β-proteins in the form of particles having diameters of approximately 10 nm to 20 nm, and these particles were designated as amylospheroid (see Hoshi, M., et al., Proc. Natl. Acad. Sci. U.S.A., 100, 6370-6375 (2003)). In accordance with such designation, a self-assembling amyloid β-protein in the form of particles having diameters of approximately 10 nm to 20 nm is referred to as “amylospheroid” in some cases in the present specification.
Amylospheroids induce neuronal cell death at a concentration equivalent to that of amyloid β-proteins that exist in the brains of patients of Alzheimer's disease, and cause phosphorylation of tau-proteins, which is another pathological marker in the process where nerves are caused to die. Since these mechanisms are consistent with the pathological conditions of Alzheimer's disease, amylospheroids were considered to be the entity of toxicity of the amyloid β-proteins in the brains. If (1) an antibody that inhibits amylospheroid formation or (2) an antibody that inhibits toxicity of amylospheroids against neuronal cells is obtained, accordingly, such an antibody can be used as a therapeutic or preventive agent for Alzheimer's disease. If (3) an antibody having a higher reactivity with amylospheroids than with amyloid precursor proteins, amyloid β monomers, or amyloid β fibrils is obtained, such an antibody can be utilized in the assay for diagnosing Alzheimer's disease.
A method for preparing an antibody that reacts with amylospheroids as an antigen is a known method indeed. Besides, rabbit polyclonal anti-amylospheroid antibodies (ASD2, ASD3), and mouse monoclonal anti-amylospheroid antibodies (MASD1, MASD2, MASD3) have been obtained already (WO 2006/016644) (hereinafter an antibody that reacts with amylospheroids is referred to as “anti-amylospheroid antibody” in some cases). However, an antibody has not yet obtained that has a low reactivity with amyloid precursor proteins, that has a specific reactivity with amylospheroids, and that inhibits the toxicity of the foregoing proteins against neuronal cells. It should be noted that the rabbit polyclonal anti-amylospheroid antibodies (ASD2, ASD3) and mouse monoclonal anti-amylospheroid antibodies (MASD1, MASD2, MASD3) disclosed in WO 2006/016644 are hereinafter referred to as the following in the present specification:    ASD2→rpASD2    ASD3→rpASD3    MASD1→mASD1    MASD2→mASD2    MASD3→mASD3    Non-Patent Document 1: Selkoe, D. J., Annu. Rev. Neurosci., 12, 463-490 (1989)    Non-Patent Document 2: Glenner, G. G. and Wong, C. W, Biochem. Biophys. Res. Commun., 120 (3), 885-890 (1984)    Non-Patent Document 3: Ihara, Y. et al., J. Biochem., 99, 1807-1810 (1986)    Non-Patent Document 4: Grundke-Iqbal, I. et al., Proc. Natl. Acad. Sci. USA., 83, 4913-4917 (1986)    Non-Patent Document 5: Lorenzo, A and Yankner, B. A, Proc. Natl. Acad. Sci. USA, 91, 12243-12247 (1994)    Non-Patent Document 6: Yankner, B. A., et. al., Science, 250, 279-282 (1990)    Non-Patent Document 7: Hoshi, M., et. al., Proc. Natl. Acad. Sci. U.S.A., 100, 6370-6375 (2003)    Patent Document 1: JP 2001-247600 A    Patent Document 2: WO2006/016644